Adjusted odds ratios were calculated using logistic regression analysis. We traced back the distribution chain of suspected food items and sampled them for microbiological analysis. Human and food isolates were sequenced using whole genome sequencing (WGS).

From October 2016 to March 2017, 54 S. Bovismorbificans cases were identified. Sequencing indicated that all were infected with identical strains. Twenty-four cases and 37 controls participated in the study. Cases were more likely to have consumed ham products than controls (aOR = 13; 95% CI: 2.0–77) and to have shopped at a supermarket chain (aOR = 7; 95% CI: 1.3–38).

Trace-back investigations led to a Belgian meat processor: one retail ham sample originating from this processor tested positive for S. Bovismorbificans and matched the outbreak strain by WGS. All ham products related to the same batch were removed from the market to prevent further cases. This investigation illustrates the importance of laboratory surveillance for all Salmonella serotypes and the usefulness of WGS in an outbreak investigation.

Outbreak of Salmonella Bovismorbificans associated with the consumption of uncooked ham products, the Netherlands, 2016 to 2017

NSW Health and NSW Food Authority have started using whole genome sequencing technology to more quickly identify a foodborne outbreak and connect it with its source, which could reduce illnesses and even deaths.

“[It’s] a significant breakthrough that could help revolutionise how food-borne illnesses are identified, understood, tracked and managed,” said Dr Craig Shadbolt, the Food Authority’s acting chief executive.

“This will be invaluable in terms of achieving the NSW Government’s Food Safety Strategy goal of reducing foodborne illnesses caused by salmonella, campylobacter and listeria by 30 per cent by 2021.”

That sounds nice, but some practical steps, like not using raw eggs in mayo, aoili, or baked Chinese ice cream, would go farther. In Australia, rates of foodborne salmonella poisoning have climbed from 38 per 100,000 people in 2004 to 76 per 100,000 in 2016, with a record-breaking 18,170 cases last year, according to the National Notifiable Diseases Surveillance System.

High-throughput whole-genome sequencing (WGS) is a revolutionary tool in public health microbiology and is gradually substituting classical typing methods in surveillance of infectious diseases. In combination with epidemiological methods, WGS is able to identify both sources and transmission-pathways during disease outbreak investigations.

This review provides the current state of knowledge on the application of WGS in the epidemiology of Campylobacter jejuni, the leading cause of bacterial gastroenteritis in the European Union.

We describe how WGS has improved surveillance and outbreak detection of C. jejuni infections and how WGS has increased our understanding of the evolutionary and epidemiological dynamics of this pathogen. However, the full implementation of this methodology in real-time is still hampered by a few hurdles. The limited insight into the genetic diversity of different lineages of C. jejuni impedes the validity of assumed genetic relationships. Furthermore, efforts are needed to reach a consensus on which analytic pipeline to use and how to define the strains cut-off value for epidemiological association while taking the needs and realities of public health microbiology in consideration.

Even so, we claim that ample evidence is available to support the benefit of integrating WGS in the monitoring of C. jejuni infections and outbreak investigations.

Use of whole-genome sequencing in the epidemiology of Campylobacter jejuni infections: state-of-knowledge

Although definitive epidemiological information demonstrating exposure to the specific recalled product was lacking, the patient reported consumption of a prepackaged romaine lettuce–containing product of either the recalled brand or a different brand.

A multinational investigation found that patient and food isolates from the recalled product were indistinguishable by pulsed-field gel electrophoresis and were highly related by whole genome sequencing, differing by four alleles by whole genome multilocus sequence typing and by five high-quality single nucleotide polymorphisms, suggesting a common source.

To our knowledge, this is the first time prepackaged lettuce has been identified as a likely source for listeriosis. This investigation highlights the power of whole genome sequencing, as well as the continued need for timely and thorough epidemiological exposure data to identify sources of foodborne infections.

Use of whole genome sequencing and patient interviews to link a case of sporadic listeriosis to consumption of prepackaged lettuce

In order to construct the history of this strain, the investigators performed whole-genome sequencing of samples of DT104 that had been collected from patients over more than 40 years, from 1969 to 2012, in 21 countries, on six continents. Very tiny changes in the genome that took place over time enabled them to construct the strain’s family tree. The sequences have also made it easy to estimate roughly when the pathogen acquired the resistance genes.

DT104’s success was due in no small part to its resistance to at least five antibiotics, including ampicillin, chloramphenicol, streptomycin, sulphonamide, and tetracycline, said corresponding author, Pimlapas Leekitcharoenphon, PhD.

Further abetting its spread, unlike other strains of DT Salmonella, DT104 was able to infect numerous livestock species, including cattle, poultry, pigs, and sheep, said Leekitcharoenphon. “Having multiple hosts increases the chances of dissemination,” she explained. Leekitcharoenphon is a postdoctoral researcher at the Research Group for Genomic Epidemiology, National Food Institute, Technical University of Denmark, Lyngby.

Using a program that took into account the rate of mutations in DT104, the investigators estimated that it first emerged in 1948 as an antibiotic susceptible pathogen. It is not clear exactly when DT104 first acquired the multidrug resistance-containing transposon. Transposons are mobile genetic elements that can carry antibiotic resistance genes, and that can jump from one genome to another. In the case of DT104, transposons have been identified as the sources of the resistance genes. The study suggests that the first acquisition of antibiotic resistance may have happened in 1972. However, multidrug-resistant DT104 was first reported in 1984 in the United Kingdom.

The new results also illuminated, for the first time, the results of a program in Denmark to eradicate all pigs infected with DT104, which had begun in 1996, but was stopped in 2000 due to financial pressures. It turns out that program was quite successful.

“If we know and understand the past, we might be able to solve the current resistance problems and prevent future ones,” said Leekitcharoenphon.

As we are approaching the twentieth anniversary of PulseNet, a network of public health and regulatory laboratories that has changed the landscape of foodborne illness surveillance through molecular subtyping, public health microbiology is undergoing another transformation brought about by so-called next-generation sequencing (NGS) technologies that have made whole-genome sequencing (WGS) of foodborne bacterial pathogens a realistic and superior alternative to traditional subtyping methods.

Routine, real-time, and widespread application of WGS in food safety and public health is on the horizon. Technological, operational, and policy challenges are still present and being addressed by an international and multidisciplinary community of researchers, public health practitioners, and other stakeholders. Expected final online publication date for the Annual Review of Food Science and Technology Volume 7 is February 28, 2016. Please seehttp://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.

Today, whole genome sequencing is all the rage, and all I can remember is, another six months of graduate school for you, Powell, go tape those gels.

Our rudimentary DNA sequencing back in 1985 involved a particular skill with masking tape so the gels wouldn’t leak.

And a lot of radioactive phosphorous.

And phenol-based extraction, which has left my one pinky finger smaller than the other.

Science.

Whole genome sequencing (WGS) has emerged as a powerful tool for comparing bacterial isolates in outbreak detection and investigation. Here, we demonstrate that WGS performed prospectively for national epidemiologic surveillance of Listeria monocytogenes has the capacity to be superior to our current approach using pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), multilocus variable-number tandem repeat analysis (MLVA), binary typing and serotyping. Initially 423 L. monocytogenes isolates underwent WGS and comparisons uncovered a diverse genetic population structure derived from three distinct lineages. MLST, binary and serotyping results inferred in silico from the WGS data were highly concordant (>99%) with laboratory typing performed in parallel. However, WGS was able to identify distinct nested clusters within groups of isolates that were otherwise indistinguishable by our current typing methods. Routine WGS was then used for prospective epidemiologic surveillance on a further 97 L. monocytogenes isolates over a 12-month period, providing a greater level of discrimination to conventional typing for inferring linkage to point source outbreaks. A risk based alert system based on WGS similarity was used to inform epidemiologists required to act on the data. Our experience shows WGS could be adopted for prospective L. monocytogenes surveillance, and investigated for other pathogens relevant to public health.

We conducted a microbiological investigation of patients and buffet items, a case–control study and a carriage study of catering staff. Isolates of Staphylococcus aureus from patients, food and catering staff were characterized and compared using traditional typing methods and whole genome sequencing.

Identical strains (sequence type ST8, spa-type t024, MLVA-type 4698, enterotoxin A FRI100) were isolated in 10 patients, shiitake mushrooms, cured ham, and in three members of staff. The case–control study strongly suggested pasta salad with pesto as the vehicle of infection (p<0.001), but this food item could not be tested, because there were no leftovers. Additional enterotoxigenic strains genetically unrelated to the outbreak strain were found in four members of staff. Non-enterotoxigenic strains with livestock-associated sequence type ST398 were isolated from three food items and two members of staff.

The main cause of the outbreak is likely to have been not maintaining the cold chain after food preparation. Whole genome sequencing resulted in phylogenetic clustering which concurred with traditional typing while simultaneously characterizing virulence and resistance traits.

Previously, samples from sick patients were sent to state and federal labs, where disease detectives ran tests to see if the infections were caused by the same bug. When enough matches emerged, typically a dozen or so, epidemiologists interviewed sick people, looking for a common food that was causing the outbreak.

But the testing wasn’t definitive, and linking one case to another took time. “While all of this was going on, more contaminated product was getting out into the public,” said Dr. Steven Musser, deputy director for scientific operations at the U.S. Food and Drug Administration’s Center for Food Safety and Applied Nutrition.

Now, the FDA is building a network of state and federal labs equipped to map out the exact DNA sequence of strains of Listeria, Salmonella and other foodborne pathogens found in sick patients. These sequences are then uploaded to a public database housed at the National Institutes of Health. The technology can not only differentiate a pathogen from multiple related species, but can also show slight mutations within the same strain.

At the same time, the FDA has begun sequencing pathogens found during routine plant inspections and adding those to the database. One benefit of that, they say, is being able to quickly connect patients within an outbreak. Another is the potential to identify the source of an outbreak after just a few patients fall ill, shortening the time it takes to get tainted food off store shelves.

To increase the odds of a match, the FDA wants manufacturers to contribute samples of pathogens found during their own plant inspections. Some contamination is common in food plants. When it is found in the manufacturing facility, but not in food products, companies generally are required only to clean it up without recalling products.

But eliminating pathogens is tough, and convincing companies to offer up potentially incriminating evidence has been a hard sell, according to interviews with public health officials, food manufacturers and experts on recalls.

The National Food Institute, Technical University of Denmark, is promoting the creation of global databases for WGS results to facilitate intelligent use of this data.

Debate in the media around foodborne outbreaks often deteriorates to simplified descriptions of unsafe foods and nobody taking action. But that is not the case. In a 2014 Salmonella enteritidis outbreak fx that affected several European countries the source was identified through the use of whole genome sequencing, WGS, in which an organism’s entire DNA-profile is mapped out. This is just one example of the work to track down and prevent people from eating unsafe food.

In August 2014 a listeria outbreak hit headlines in Denmark. The outbreak had broken out in November 2013 and went on for months making at least 38 people sick and claiming 16 lives. The fact that this event was recognized as an outbreak stems partly from the fact that Danish labs used WGS technology to compare the listeria isolates from humans with bacterial strains isolated from food, thereby enabling linking the outbreak to a food source.

The use of WGS technology is making it possible for scientists to more quickly link many more cases of foodborne disease to an outbreak. The National Food Institute is working to make WGS data widely available through the creation of global databases for WGS results.

The use of WGS in solving foodborne outbreaks is described in greater detail in two articles in Pan European Networks Science & Technology: Foodborne outbreaks, not the whole story and Microbiology 2.0. The article is written by Jørgen Schlundt, former director of the National Food Institute, now professor at DTU Management Engineering.

The National Food Institute is part of the Global Microbial Identifier network, which works to create a global system of DNA genome databases that can be used for microbial and infectious disease identification and diagnostics. The databases will enable the identification of relevant genes and the comparison of genomes to outbreaks and emerging pathogens.